Project Summary Genetic polymorphisms have recently emerged as risk factors for neurocognitive disorder (NCD) following anesthesia and surgery, but there is a gap in our understanding whether or how synaptic dysfunction plays a role in these cognitive deficits. Such knowledge is critical to assessing the risk of necessary medical procedures that require anesthesia, particularly in vulnerable populations. The long-term goal of this proposal is to define the cellular and molecular mechanisms of sustained general anesthetic actions on synaptic plasticity. The overall objective in this application is to identify signaling pathways involved in persistent structural dendritic spine changes following anesthetic exposure. Dendritic spines are postsynaptic structures on mature synapses that are critical for learning and memory and are associated with cognitive and developmental dysfunction in multiple neurological disorders. The central hypothesis is that general anesthetics reduce brain-derived neurotrophic factor (BDNF) release, resulting in impaired presynaptic function, alterations in dendritic spine structure, and deficits in neuronal activity and spatial learning and memory. This hypothesis has been formulated on the basis of preliminary data obtained in the applicant?s laboratory. The rationale for the proposed research is that, once it is known how anesthetics induce permanent spine loss, pharmacological manipulation of these molecular targets during anesthesia or design of new anesthetic agents that avoid these effects will be possible. Guided by compelling preliminary data, this hypothesis will be tested by three specific aims: 1) Identify the role of BDNF in reducing synaptic vesicle (SV) exocytosis by volatile anesthetics; 2) Define the role of BDNF in volatile anesthetic- induced changes on dendritic spine density and morphology; and 3) Elucidate the role of BDNF in isoflurane- induced effects on hippocampal-dependent synaptic plasticity. For Aim 1, proven optogenetic biosensors (BDNF- pH, vGlut1-pH and the Ca2+ indicator GCaMP6), already established as feasible in the applicant?s hands, will be used to test anesthetic effects on BDNF release and the impact of these changes on inhibition of Ca2+ entry and SV exocytosis. For Aim 2, structural spine changes due to reduced BDNF will be investigated in dissociated hippocampal cultures and intact brain slices using time-lapse fluorescence microscopy and Golgi staining, respectively. For Aim 3, the functional outcome of reduced BDNF signaling by isoflurane on real-time neuronal activity corresponding to impairments in learning and memory will be assessed with fiber photometry and behavioral testing. These functional and structural studies will be conducted with or using neurons or slices from transgenic mice with reduced BDNF secretion due to a common human polymorphism (Val66Met) recently identified as a risk factor for NCD. This model provides an innovative approach to elucidate mechanisms underlying anesthetic-induced pre- and post-synaptic dysfunction associated with impairments in learning and memory. Prevention of long-term synaptic deficits following anesthesia in at-risk individuals have broad translational importance to preoperative risk assessment and anesthesia recovery.